Molecular detection of Anaplasma Capra and Anaplasma marginale in Rhipicephalus microplus ticks infesting cows

Ticks are the second most significant hematophagous ectoparasites, serving as vectors for numerous infectious diseases worldwide, surpassed only by mosquitoes. This study aimed to molecularly characterize ticks infesting cows and their associated pathogens in Khyber Pakhtunkhwa, Pakistan, with a focus on assessing the potential risk of zoonotic disease transmission. Between January 2023 and June 2024, 240 cows from six districts: Buner, Swat, Lower Dir, Upper Dir, Shangla, and Chitral, were examined for tick infestations. DNA was extracted using the phenol-chloroform method, and pathogen detection was performed via PCR targeting the gltA gene, while tick species were confirmed by amplifying partial 12 S rDNA sequences. The study revealed an average tick intensity of 12.73 ticks per infested cow and a mean abundance of 10.45 ticks per examined cow. In total, 2,507 ticks were collected and morphologically identified as Rhipicephalus microplus, comprising 915 females, 801 males, 510 nymphs, and 281 larvae. The highest tick burdens were observed in Buner and Swat, followed by Lower Dir, Upper Dir, and Shangla, with Chitral showing the lowest prevalence. Molecular screening detected multiple pathogens, notably the human pathogen Anaplasma capra and the cow pathogen Anaplasma marginale; co-infections were occasionally observed. This paper presents the first phenotypic and phylogenetic characterization of A. capra in ticks from cows in Pakistan, emphasizing the zoonotic transmission risk. These findings advance our understanding of R. microplus associated pathogen risks and underscore the need for enhanced surveillance in the region.